The cranial apparatus glands of the canthariphilous Pyrochroa coccinea (Coleoptera: Pyrochroidae: Pyrochroinae), and their implications in sexual behaviour.

Some Pyrochroidae species are known as "canthariphilous" for their attraction to cantharidin (CTD), a toxic terpene with anti-predatory effects, produced in nature by only two beetle families (Meloidae and Oedemeridae). It has been demonstrated that males of Neopyrochroa flabellata ingesting CTD are positively selected by females. Indeed, the compound is re-emitted from a glandular cranial apparatus as secretions that are licked up by females during courtship behaviour, inducing copulation. Herein, we provide the first description of the glands associated to the cranial apparatus of male Pyrochroinae using the European species Pyrochroa coccinea as a model. Morphological analyses show that the cranial apparatus consists of a concave pit lined with short setae retaining secretions emitted through numerous glandular pores. Ultrastructural investigations reveal the presence of two different class 3 glands (Gl.A and Gl.B), intermixed at the level of the pit but exhibiting distinct features. Gl.A are mainly characterised by short conducting canals, rounded nuclei and electrondense vesicles while Gl.B are characterised by long conducting canals, irregular nuclei, vesicles containing a particulate substance and a multifolded plasma membrane. Observations of sexual behaviour are also reported for P. coccinea and compared to N. flabellata, confirming the involvement of cranial apparatus secretions in courtship behaviour.

Larval features of the Italian endemic Pyrochroa serraticornis kiesenwetteri Fairmaire, 1849 (Coleoptera: Pyrochroidae: Pyrochroinae) solve taxonomic uncertainties.

Recent findings rearranged the taxonomy of the European species of the genus Pyrochroa Geoffroy (Coleoptera: Pyrochroidae), and to date three species are recognized: P. bifoveata Molfini et al., 2022 and P. coccinea (Linnaeus, 1761) with cryptic adults and distinctive larvae, and the polytypic P. serraticornis (Scopoli, 1763), including the subspecies kiesenwetteri Fairmaire, 1849. Incongruences between molecular and morphological analyses questioned the recognition of P. s. serraticornis and P. s. kiesenwetteri as taxa of the same species. In the present paper, observations of larval characters of P. s. kiesenwetteri confirm this taxon as a subspecies of P. serraticornis. Moreover, new characters of P. s. serraticornis larvae are offered by analysing specimens from a new European locality.

New Evidence of Canthariphily: Tilloidea transversalis (Coleoptera: Cleridae) Sequestering Cantharidin From Lydus trimaculatus (Coleoptera: Meloidae).

Cantharidin (CTD) is a defensive compound autogenously and exclusively produced by two phylogenetically related beetle families: Meloidae and Oedemeridae. Although this molecule usually acts as a strong deterrent against potential predators and parasites, some arthropod species, collectively named 'canthariphilous species', are attracted to CTD. Some species can sequester CTD from the CTD-producing species, using it as a chemical defense against enemies. The present paper focuses on the first-ever description of canthariphilous interactions between a checkered beetle species (Coleoptera: Cleridae) and a CTD -producing species. Field observations revealed individuals of the phytophagous beetle Tilloidea transversalis (Charpentier, 1825) (Coleoptera: Cleridae) biting individuals of the blister beetle Lydus trimaculatus (Fabricius, 1775) (Coleoptera: Meloidae). Laboratory behavioral experiments followed to verify if this peculiar behavior of T. transversalis also occurs on other co-occurring species. Moreover, chemical analyses were performed to assess whether T. transversalis can sequester CTD. Our results show that T. transversalis only attacks CTD-producing species. However, while chemical analyses prove that T. transversalis can sequester CTD from the hemolymph of L. trimaculatus, some clues (based on a CTD-baited traps sampling) suggest that this beetle, contrarily to other canthariphilous species, does not appear to show a high attraction to pure synthetic CTD. Thus, other unknown signals, alone or in combination with CTD, could be implicated in triggering the canthariphilous behaviors of T. transversalis.

Unraveling the role of male reproductive tract and haemolymph in cantharidin-exuding Lydus trimaculatus and Mylabris variabilis (Coleoptera: Meloidae): a comparative transcriptomics approach.

BACKGROUND: Meloidae (blister beetles) are known to synthetize cantharidin (CA), a toxic and defensive terpene mainly stored in male accessory glands (MAG) and emitted outward through reflex-bleeding. Recent progresses in understanding CA biosynthesis and production organ(s) in Meloidae have been made, but the way in which self-protection is achieved from the hazardous accumulation and release of CA in blister beetles has been experimentally neglected. To provide hints on this pending question, a comparative de novo assembly transcriptomic approach was performed by targeting two tissues where CA is largely accumulated and regularly circulates in Meloidae: the male reproductive tract (MRT) and the haemolymph. Differential gene expression profiles in these tissues were examined in two blister beetle species, Lydus trimaculatus (Fabricius, 1775) (tribe Lyttini) and Mylabris variabilis (Pallas, 1781) (tribe Mylabrini). Upregulated transcripts were compared between the two species to identify conserved genes possibly involved in CA detoxification and transport. RESULTS: Based on our results, we hypothesize that, to avoid auto-intoxication, ABC, MFS or other solute transporters might sequester purported glycosylated CA precursors into MAG, and lipocalins could bind CA and mitigate its reactivity when released into the haemolymph during the autohaemorrhaging response. We also found an over-representation in haemolymph of protein-domains related to coagulation and integument repairing mechanisms that likely reflects the need to limit fluid loss during reflex-bleeding. CONCLUSIONS: The de novo assembled transcriptomes of L. trimaculatus and M. variabilis here provided represent valuable genetic resources to further explore the mechanisms employed to cope with toxicity of CA in blister beetle tissues. These, if revealed, might help conceiving safe and effective drug-delivery approaches to enhance the use of CA in medicine.

Larval features illuminating adult taxonomy? Case study in the European cardinal beetle species of the genus Pyrochroa (Coleoptera: Pyrochroidae: Pyrochroinae).

Larvae of the two European species of the genus Pyrochroa (P. coccinea and P. serraticornis) are redescribed, for the first time based on Italian samples, and new relevant and distinctive characters have emerged from P. serraticornis. Particularly, the presence of a single urogomphal pit between urogomphi (representing a new condition for the entire family), and new characters of mandibles, open new questions about the current taxonomic status of the European species.

Comparative morphology of myrmecophilous immature stages of European Microdon species (Diptera: Syrphidae): updated identification key and new diagnostic characters.

Hoverflies (Diptera: Syrphidae) of the genus Microdon Meigen have larvae that live in ant nests where they are predatory on ant larvae. Reflecting the exceptional challenges of this very specialized lifestyle, Microdon eggs, larvae and puparia are highly distinctive in their morphology. Detailed descriptions of these immature stages is, however, lacking for all but a very few species, and much of this has been limited through the sole use of light microscopes. Here, using Scanning Electron Microscopy (SEM), we present detailed, comparative descriptions of the immature stages of three European Microdon species: M. analis, M. devius and M. myrmicae. Given that many adult Microdon species are very similar to each other in their outward appearance, we demonstrate that the morphology of their immature stages can improve our understanding of the phylogeny of the genus. We also discuss how particular adaptations of the immature morphology may allow their myrmecophilous life within ant nests. In this paper new diagnostic features are also presented to distinguish M. myrmicae from its sibling species M. mutabilis-the two are morphologically indistinguishable as adults.